Components and proteolytic processing sites of arylsulfatase B from human placenta

Area of Science:

• Biochemistry
• Molecular Biology
• Enzymology

Background:

• Mature arylsulfatase B from human sources typically consists of 43 kDa and 8 kDa chains.
• Previous characterization did not identify all components of the mature enzyme.

Purpose of the Study:

• To identify and characterize all components of human arylsulfatase B purified from placenta.
• To elucidate the post-translational processing and maturation of human arylsulfatase B.

Main Methods:

• Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using Tris-Tricine system.
• Carbohydrate staining (periodic acid-Schiff), lectin blotting (Con-A), and enzymatic treatments (endo-glycosidase, phosphorylation).
• N-terminal sequencing of isolated polypeptide components.

Main Results:

• A 7 kDa component, containing a carbohydrate moiety, was identified in addition to the previously known 43 kDa (carbohydrate-containing) and 8 kDa (non-carbohydrate-containing) chains.
• All three chains are linked by disulfide bonds, appearing as a single 58 kDa polypeptide under non-reducing conditions.
• N-terminal sequencing revealed distinct starting points for each component (Ala-41, Ala-424, Asp-466), indicating proteolytic cleavage sites.

Conclusions:

• Human arylsulfatase B undergoes significant proteolytic processing during maturation, yielding at least three distinct chains.
• The identified components originate from different positions within the precursor protein, suggesting multiple cleavage events after signal peptide removal.
• The presence of a glycosylated 7 kDa chain highlights the complexity of arylsulfatase B post-translational modification.